Strontium promotes osteogenic differentiation by activating autophagy via the the AMPK/mTOR signaling pathway in MC3T3‑E1 cells.

Abstract

Strontium (Sr) is an alkaline earth metal that exerts the dual effect of improving bone formation and suppressing bone resorption, resulting in increased bone apposition rates and bone mineral density. However, the mechanisms through which Sr exerts these beneficial effects on bone have yet to be fully elucidated. The present study aimed to reveal the underlying molecular mechanisms associated with Sr-induced osteogenic differentiation. The effects of Sr on cell proliferation and osteogenic differentiation were analyzed by MTT assay, RT-qPCR, western blot analysis, alkaline phosphatase (ALP) and Alizarin red staining assays. The extent of autophagy was determined by monodansylca-daverine (MDC) staining and western blot analysis of two markers of cellular autophagic activity, the steatosis-associated protein, sequestosome-1 (SQSTM1/p62), and the two isoforms of microtubule-associated protein 1 light chain 3 (LC3), LC-3-I/II. The expression levels of AMP-activated protein kinase (AMPK) and mammalian target of rapamycin (mTOR) were also detected by western blot analysis. Sr at a concentration of 3 mM exerted the most pronounced effect on osteogenic differentiation, without any apparent cell toxicity. At the same time, cellular autophagy was active during this process. Subsequently, autophagy was blocked by 3-methyl-adenine, and the enhancement of osteogenic differentiation in response to Sr was abrogated. Additionally, the phosphorylation level of AMPK was significantly increased, whereas that of mTOR was significantly decreased, in the Sr-treated group. Taken together, the findings of the present study demonstrate that Sr stimulates AMPK-activated autophagy to induce the osteogenic differentiation of MC3T3-E1 cells.